Exposure to elevated indoor levels of nitrogen dioxide (NO2) represent a significant health problem, most notably increased airway reactivity. Alterations of the airway epithelial barrier and active ion transport pathways play major roles in increased reactivity. We found that NO2 caused an enhancement of Na-K-ATPase (sodium pump) activity that preceded a significant decline of transepithelial resistance (Rte). We hypothesize that NO2 exposure may enhance Na-K-ATPase activity through an interplay of two mechanisms: first, increased Na entry caused by either oxidative damage of apical Na conductive pathways or a rise of intracellular Ca levels followed by subsequent Na/Ca exchange; and, secondly, by altered phosphorylation of the pump. Potentially, decreased Rte and enhanced pump activity are indirectly linked by altered phosphorylation. We further hypothesize that these effects are mediated by lipid peroxidation products. In Aim #1, we will examine the effects of NO2 or lipid peroxidation products on airway epithelial Na entry and its relationship to alterations of apical Na conductive pathways and intracellular Ca levels. The effects of increased Na entry on pump activity and its cellular distribution will be examined. In Aim #2, we will examine if decreased Rte and enhanced Na/K-ATPase activity are indirectly linked through alterations of signal transduction pathways. We will determine if NO2 or lipid peroxidation products alter phosphorylation of the tight junctional protein, zonula occludens-1 (ZO1) and the sodium pump alpha subunit. If alterations in phosphorylation are found, we will use inhibitors of protein kinase and phosphatase activities to identify second messenger pathways that mediate these changes. We will attempt to establish a correlation between increased ZO1 phosphorylation and decreased Rte. In Aim #3, based upon results in Aim #2, we will examine the effects of NO2 or lipid peroxidation products on the distribution of cellular activity and abundance for selected protein kinases and phosphatases in membrane and cytosol. These studies will provide a greater mechanistic understanding of sublethal injury resulting from low level NO2 exposure and airway epithelial compensatory responses.